2-Benzyl-3-arylbenzothiophenes

ABSTRACT

The present invention provides a compound of formula I ##STR1## wherein R 1  is hydrogen, a hydroxy protecting group, or methyl; 
     R 2  and R 3  are independently C 1  -C 6  alkyl, or R 2  and R 3 , together with the nitrogen to which they are attached, form a heterocycle selected from the group consisting of pyrrolidino, hexamethyleneimino, morpholino, and piperidino; 
     Z is bromo, chloro, fluoro, hydroxy, a protected hydroxy, or hydrogen; and pharmaceutically acceptable salts and solvates thereof. 
     The present invention further provides pharmaceutical compositions containing compounds of formula I, optionally containing estrogen or progestin, and the use of such compounds alone, or in combination with estrogen or progestin, for alleviating the symptoms of post-menopausal syndrome, particularly osteoporosis, cardiovascular related pathological conditions, and estrogen-dependent cancer. 
     The compounds of the present invention also are useful for inhibiting uterine fibroid disease and endometriosis in women and aortal smooth muscle cell proliferation, particularly restenosis, in humans. 
     Also encompassed are novel intermediates.

This application is a division, of application Ser. No. 08/396,021 filedFeb. 28, 1995, U.S. Pat. No. 5,552,401.

FIELD OF THE INVENTION

This invention relates to the fields of pharmaceutical and organicchemistry and provides novel benzothiophene compounds which are usefulfor the treatment of the various medical indications associated withpost-menopausal syndrome, and uterine fibroid disease, endometriosis,and aortal smooth muscle cell proliferation.

BACKGROUND OF THE INVENTION

"Post-menopausal syndrome" is a term used to describe variouspathological conditions which frequently affect women who have enteredinto or completed the physiological metamorphosis known as menopause.Although numerous pathologies are contemplated by the use of this term,three major effects of post-menopausal syndrome are the source of thegreatest long-term medical concern: osteoporosis, cardiovascular effectssuch as hyperlipidemia, and estrogen-dependent cancer, particularlybreast and uterine cancer.

Osteoporosis describes a group of diseases which arise from diverseetiologies, but which are characterized by the net loss of bone mass perunit volume. The consequence of this loss of bone mass and resultingbone fracture is the failure of the skeleton to provide adequatestructural support for the body. One of the most common types ofosteoporosis is that associated with menopause. Most women lose fromabout 20% to about 60% of the bone mass in the trabecular compartment ofthe bone within 3 to 6 years after the cessation of mensus. This rapidloss is generally associated with an increase of bone resorption andformation. However, the resorptive cycle is more dominant and the resultis a net loss of bone mass. Osteoporosis is a common and serious diseaseamong post-menopausal women.

There are an estimated 25 million women in the United States, alone, whoare afflicted with this disease. The results of osteoporosis arepersonally harmful and also account for a large economic loss due itschronicity and the need for extensive and long term support(hospitalization and nursing home care) from the disease sequelae. Thisis especially true in more elderly patients. Additionally, althoughosteoporosis is not generally thought of as a life threateningcondition, a 20% to 30% mortality rate is related with hip fractures inelderly women. A large percentage of this mortality rate can be directlyassociated with post-menopausal osteoporosis.

The most vulnerable tissue in the bone to the effects of post-menopausalosteoporosis is the trabecular bone. This tissue is often referred to asspongy or cancellous bone and is particularly concentrated near the endsof the bone (near the joints) and in the vertebrae of the spine. Thetrabecular tissue is characterized by small osteoid structures whichinterconnect with each other, as well as the more solid and densecortical tissue which makes up the outer surface and central shaft ofthe bone. This inter-connected network of trabeculae gives lateralsupport to the outer cortical structure and is critical to thebio-mechanical strength of the overall structure. In post-menopausalosteoporosis, it is, primarily, the net resorption and loss of thetrabeculae which leads to the failure and fracture of bone. In light ofthe loss of the trabeculae in post-menopausal women, it is notsurprising that the most common fractures are those associated withbones which are highly dependent on trabecular support, e.g., thevertebrae, the neck of the weight bearing bones such as the femur andthe fore-arm. Indeed, hip fracture, collies fractures, and vertebralcrush fractures are hall-marks of post-menopausal osteoporosis.

At this time, the only generally accepted method for treatment ofpost-menopausal osteoporosis is estrogen replacement therapy. Althoughtherapy is generally successful, patient compliance with the therapy islow primarily because estrogen treatment frequently produces undesirableside effects.

Throughout premenopausal time, most women have less incidence ofcardiovascular disease than age-matched men. Following menopause,however, the rate of cardiovascular disease in women slowly increases tomatch the rate seen in men. This loss of protection has been linked tothe loss of estrogen and, in particular, to the loss of estrogen'sability to regulate the levels of serum lipids. The nature of estrogen'sability to regulate serum lipids is not well understood, but evidence todate indicates that estrogen can upregulate the low density lipid (LDL)receptors in the liver to remove excess cholesterol. Additionally,estrogen appears to have some effect on the biosynthesis of cholesterol,and other beneficial effects on cardiovascular health.

It has been reported in the literature that post-menopausal women havingestrogen replacement therapy have a return of serum lipid levels toconcentrations to those of the pre-menopausal state. Thus, estrogenwould appear to be a reasonable treatment for this condition. However,the side-effects of estrogen replacement therapy are not acceptable tomany women, thus limiting the use of this therapy. An ideal therapy forthis condition would be an agent which would regulate the serum lipidlevel as does estrogen, but would be devoid of the side-effects andrisks associated with estrogen therapy.

The third major pathology associated with post-menopausal syndrome isestrogen-dependent breast cancer and, to a lesser extent,estrogen-dependent cancers of other organs, particularly the uterus.Although such neoplasms are not solely limited to a post-menopausalwomen, they are more prevalent in the older, post-menopausal population.Current chemotherapy of these cancers has relied heavily on the use ofanti-estrogen compounds such as, for example, tamoxifen. Although suchmixed agonist-antagonists have beneficial effects in the treatment ofthese cancers, and the estrogenic side-effects are tolerable in acutelife-threatening situations, they are not ideal. For example, theseagents may have stimulatory effects on certain cancer cell populationsin the uterus due to their estrogenic (agonist) properties and they may,therefore, be contraproductive in some cases. A better therapy for thetreatment of these cancers would be an agent which is an anti-estrogencompound having negligible or no estrogen agonist properties onreproductive tissues.

In response to the clear need for new pharmaceutical agents which arecapable of alleviating the symptoms of, inter alia, post-menopausalsyndrome, the present invention provides new benzothiophene compounds,pharmaceutical compositions thereof, and methods of using such compoundsfor the treatment of post-menopausal syndrome and other estrogen-relatedpathological conditions such as those mentioned below.

Uterine fibrosis (uterine fibroid disease) is an old and ever presentclinical problem which goes under a variety of names, including uterinefibroid disease, uterine hypertrophy, uterine lieomyomata, myometrialhypertrophy, fibrosis uteri, and fibrotic metritis. Essentially, uterinefibrosis is a condition where there is an inappropriate deposition offibroid tissue on the wall of the uterus.

This condition is a cause of dysmenorrhea and infertility in women. Theexact cause of this condition is poorly understood but evidence suggeststhat it is an inappropriate response of fibroid tissue to estrogen. Sucha condition has been produced in rabbits by daily administrations ofestrogen for 3 months. In guinea pigs, the condition has been producedby daily administration of estrogen for four months. Further, in rats,estrogen causes similar hypertrophy.

The most common treatment of uterine fibrosis involves surgicalprocedures both costly and sometimes a source of complications such asthe formation of abdominal adhesions and infections. In some patients,initial surgery is only a temporary treatment and the fibroids regrow.In those cases a hysterectomy is performed which effectively ends thefibroids but also the reproductive life of the patient. Also,gonadotropin releasing hormone antagonists may be administered, yettheir use is tempered by the fact they can lead to osteoporosis. Thus,there exists a need for new methods for treating uterine fibrosis, andthe methods of the present invention satisfy that need.

Endometriosis is a condition of severe dysmenorrhea, which isaccompanied by severe pain, bleeding into the endometrial masses orperitoneal cavity and often leads to infertility. The cause of thesymptoms of this condition appear to be ectopic endometrial growthswhich respond inappropriately to normal hormonal control and are locatedin inappropriate tissues. Because of the inappropriate locations forendometrial growth, the tissue seems to initiate local inflammatory-likeresponses causing macrophage infiltration and a cascade of eventsleading to initiation of the painful response. The exact etiology ofthis disease is not well understood and its treatment by hormonaltherapy is diverse, poorly defined, and marked by numerous unwanted andperhaps dangerous side effects.

One of the treatments for this disease is the use of low dose estrogento suppress endometrial growth through a negative feedback effect oncentral gonadotropin release and subsequent ovarian production ofestrogen; however, it is sometimes necessary to use continuous estrogento control the symptoms. This use of estrogen can often lead toundesirable side effects and even the risk of endometrial cancer.

Another treatment consists of continuous administration of progestinswhich induces amenorrhea and by suppressing ovarian estrogen productioncan cause regressions of the endometrial growths. The use of chronicprogestin therapy is often accompanied by the unpleasant CNS sideeffects of progestins and often leads to infertility due to suppressionof ovarian function.

A third treatment consists of the administration of weak androgens,which are effective in controlling the endometriosis; however, theyinduce severe masculinizing effects. Several of these treatments forendometriosis have also been implicated in causing a mild degree of boneloss with continued therapy. Therefore, new methods of treatingendometriosis are desirable.

Smooth aortal muscle cell proliferation plays an important role indiseases such as atherosclerosis and restenosis. Vascular restenosisafter percutaneous transluminal coronary angioplasty (PTCA) has beenshown to be a tissue response characterized by an early and late phase.The early phase occurring hours to days after PTCA is due to thrombosiswith some vasospasms while the late phase appears to be dominated byexcessive proliferation and migration of aortal smooth muscle cells. Inthis disease, the increased cell motility and colonization by suchmuscle cells and macrophages contribute significantly to thepathogenesis of the disease. The excessive proliferation and migrationof vascular aortal smooth muscle cells may be the primary mechanism tothe reocclusion of coronary arteries following PTCA, atherectomy, laserangioplasty and arterial bypass graft surgery. See "IntimalProliferation of Smooth Muscle Cells as an Explanation for RecurrentCoronary Artery Stenosis after Percutaneous Transluminal CoronaryAngioplasty," Austin et al., Journal of the American College ofCardiology, 8: 369-375 (August 1985).

Vascular restenosis remains a major long term complication followingsurgical intervention of blocked arteries by percutaneous transluminalcoronary angioplasty (PTCA), atherectomy, laser angioplasty and arterialbypass graft surgery. In about 35% of the patients who undergo PTCA,reocclusion occurs within three to six months after the procedure. Thecurrent strategies for treating vascular restenosis include mechanicalintervention by devices such as stents or pharmacologic therapiesincluding heparin, low molecular weight heparin, coumarin, aspirin, fishoil, calcium antagonist, steroids, and prostacyclin. These strategieshave failed to curb the reocclusion rate and have been ineffective forthe treatment and prevention of vascular restenosis. See "Prevention ofRestenosis after Percutaneous Transluminal Coronary Angioplasty: TheSearch for a `Magic Bullet`," Hermans et al., American Heart Journal,122: 171-187 (July 1991).

In the pathogenesis of restenosis excessive cell proliferation andmigration occurs as a result of growth factors produced by cellularconstituents in the blood and the damaged arterial vessel wall whichmediate the proliferation of smooth muscle cells in vascular restenosis.

Agents that inhibit the proliferation and/or migration of smooth aortalmuscle cells are useful in the treatment and prevention of restenosis.The present invention provides for the use of compounds as smooth aortalmuscle cell proliferation inhibitors and, thus inhibitors of restenosis.

SUMMARY OF THE INVENTION

The present invention relates to compounds of formula I ##STR2## whereinR₁ is hydrogen, a hydroxy protecting group, or methyl;

R₂ and R₃ are independently C₁ -C₆ alkyl, or R₂ and R₃, together withthe nitrogen to which they are attached, form a heterocycle selectedfrom the group consisting of pyrollidino, hexamethyleneimino,morpholino, and piperidino;

Z is bromo, chloro, fluoro, hydroxy, protected hydroxy, or hydrogen; andpharmaceutically acceptable salts and solvates thereof.

The present invention further relates to pharmaceutical compositionscontaining compounds of formula I, optionally containing estrogen orprogestin, and the use of such compounds, alone, or in combination withestrogen or progestin, for alleviating the symptoms of post-menopausalsyndrome, particularly osteoporosis, cardiovascular related pathologicalconditions, and estrogen-dependent cancer. As used herein, the term"estrogen" includes steroidal compounds having estrogenic activity suchas, for example, 17β-estradiol, estrone, conjugated estrogen(Premarin®), equine estrogens, 17β-ethynyl estradiol, and the like. Asused herein, the term "progestin" includes compounds havingprogestational activity such as, for example, progesterone,norethylnodrel, nongestrel, megestrol acetate, norethindrone, and thelike.

The compounds of the present invention also are useful for inhibitinguterine fibroid disease and endometriosis in women and aortal smoothmuscle cell proliferation, particularly restenosis, in humans.

Also provided are novel intermediates.

DETAILED DESCRIPTION OF THE INVENTION

One aspect of the present invention includes compounds of formula I##STR3## wherein R₁ is hydrogen, a hydroxy protecting group, or methyl;

R₂ and R₃ are independently C₁ -C₆ alkyl, or R₂ and R₃, together withthe nitrogen to which they are attached, form a heterocycle selectedfrom the group consisting of pyrollidino, hexamethyleneimino,morpholino, and piperidino;

Z is bromo, chloro, fluoro, hydroxy, a protected hydroxy or hydrogen;and pharmaceutically acceptable salts and solvates thereof.

The invention also encompasses compounds of the formula (XIII) ##STR4##wherein R_(1a) is hydroxy, protected hydroxy, or methoxy;

Z' is bromo, chloro, fluoro, hydrogen, hydroxy, or protected hydroxy;and

R₄ is hydroxy, protected hydroxy, or a group of the formula ##STR5##wherein R₂ and R₃ are independently C₁ -C₆ alkyl, or together with thenitrogen to which they are attached, form a heterocycle selected fromthe group consisting of pyrollidino, hexamethyleneimino, morpholino, andpiperidino; and salts and solvates thereof.

General terms used in the description of compounds herein described beartheir usual meanings. For example, "C₁ -C₆ alkyl" refers to straight orbranched aliphatic chains of 1 to 6 carbon atoms including methyl,ethyl, propyl, isopropyl, butyl, n-butyl, pentyl, isopentyl, hexyl,isohexyl, and the like.

The compounds of the invention may be prepared by methodology known inthe art. For example, a 3-keto compound of the formula ##STR6## may bereacted with aryllithiumbromide (or aryl magnesium bromide) having theformula (III), ##STR7## The reaction is desirably carried out in anunreactive solvent, such as tetrahydrofuran or ether. The reactionproduct is dehydrated by heating with hydrochloric acid in ethanol.

The product of the reaction may be isolated directly as thehydrochloride salt, or employing basification, as the free base.Further, the free base may employ a further salt formation to provide anacid-addition salt.

The compounds of formula (II) may be prepared by hydrogenation of thecorresponding compounds of the formula (IV) ##STR8## in the presence ofa catalyst, such as palladium or platinum oxide, at room temperature.Suitable solvents for the reaction include alcohols and ethyl acetate.

Another example of synthesis involves reacting a compound of the formula(V) ##STR9## with magnesium bromide etherate, followed by treatment withlithium aluminum hydride. The compounds of formula (V) may be preparedby treating compounds of formula (IV) with aryllithium bromide (or arylmagnesium bromide) having the formula (III).

The compounds of formula (IV) may be synthesized in an analogous manneras set out in U.S. Pat. No. 5,354,861. A 3-keto-benzothiophene compoundof formula (VI) ##STR10## may be condensed with the appropriate aldehydein catalytic amounts of hydrochloric acid.

Another method of preparation includes using an analogous method asdescribed in Jones et al., U.S. Pat. No. 4,075,227, incorporated hereinby reference. A thiol of the formula VII ##STR11## wherein R_(1a) is aprotected-hydroxy, is reacted with an α-haloacetophenone of the formula(VIII) ##STR12## where R₄ is protected hydroxy, and Y is halo, toproduce the compound of formula (IX) ##STR13## Compound IX isring-closed in the presence of an acid, such as aryl sulfonic acid, analkane sulfonic acid, sulfuric acid, polyphosphoric acid, or the like,to form a compound of formula (X) ##STR14## Compound X is then reactedin the presence of a Friedel-Crafts catalyst, such as aluminum chloride,with a benzoyl chloride of the formula (XI) ##STR15## wherein Z' is Z,or protected hydroxy, to produce ##STR16## Compound XII is then placedin an appropriate solvent such as diethyl ether, THF, dioxane, or thelike. Thereafter, Compound XII is reacted with a reducing agent, such aslithium aluminum hydride, (LAH), under an inert gas, such as nitrogen.The amount of reducing agent used is an amount sufficient to reduce thecarbonyl to the carbinol of formula (XIII) ##STR17## This intermediateis novel, as well as when R₄ is ##STR18## or when R_(1a) is alsohydroxy, or protected hydroxy, Z' is also hydroxy, and R₄ is alsohydroxy. Thereafter, Compound XIII may be further reduced to provide the2-methylene linker. This may be accomplished by placing Compound XIII inan appropriate solvent, and cooled under an inert gas. To this is addeda suitable trialkylsilane reducing agent, preferably triethylsilyl, anda reasonably strong protic acid such as hydrochloric acid,trifluoracetic acid, or the like. Suitable solvents includedichloromethane, 1,2 dichloroethene, and chlorobenzene. The temperaturefor this reaction is between -5° C. to about 20° C. If R₁ and/or Z ishydroxy, R_(1a) and Z' are protected hydroxys during the reaction, andthe hydroxy protecting group (s) removed via known methodology.

To convert R₄ to ##STR19## either prior to the reduction to themethylene or afterwards, any other hydroxy should be protected, and thecompound (either XIII, or I, with any hydroxys protected) is treatedwith a moderately strong base, such as sodium hydroxide, in the presenceof a compound of formula ##STR20## in which Q is halo, such as bromo orchloro.

Hydroxy protecting groups suitable for the above are known in the art.Preferred protecting groups include C₁ -C₄ alkyl. See e.g., Barkin,"Protective Groups in Organic Chemistry," J. G. W. McOmie (ed.), PlenumPass, NY, N.Y., 1973, Chapter 2, and Greene, "Protective Groups inOrganic Synthesis," John Wiley and Sons, NY, N.Y., 1981, Chapter 7, incorporated herein by reference.

The following examples are intended to illustrate the inventiondescribed herein without unduly restricting it.

EXAMPLE 12-(p-Chlorobenzyl-3-[p-(2-dimethylaminoethoxy)phenyl]-6-methoxybenzo[b]thiophene)

n-Butyl lithium (5.27 ml, 7.80 mole) in THF is added dropwise to asolution of 4-[2-dimethylaminoethoxy]phenyl bromide (1.90 g, 7.80 mmol)in THF (5 ml) at -78° C. After stirring for 30 minutes at -78° C.,2-(p-chlorobenzyl)-6-methoxy-3-keto-(2H)benzothiophene (1.58 g, 5.20mmol) in THF is added dropwise to the mixture at -78° C. The reactionmixture is stirred at -78° C. for 1 hour, and allowed to warm to roomtemperature. After stirring for 18 hours, it is quenched with saturatedammonium chloride and THF is evaporated off. The residue is dissolved inethyl acetate (30 ml), washed with sodium chloride (2×20 ml), dried(anhydrous magnesium sulphate) and the solvent evaporated off. Theremaining material is purified by flash chromatography (eluent,dichloromethane:hexane:triethylamine=1:1:0.2). The alcohol withoutcharacterization is dehydrated. A solution of methanol (0.40 ml)saturated with dry hydrogen chloride gas is added to the tertiaryalcohol dissolved in sodium-dried ether (40 ml). The product is formedand is filtered and recrystallized from ethanol to give thehydrochloride salt of the title compound.

EXAMPLE 22-(p-Chlorobenzyl)-3-[p-(2-(1-pyrollidino)ethoxy)phenyl]-6-methoxybenzo[b]thiophene

4-[2-(1-Pyrollidino)ethoxy]phenyl bromide (1.41 g, 5.20 mmol) in dry THF(50 ml) is added to magnesium turnings (312 mg, 13.0 mmol) in a dry3-necked round bottom flask under an oxygen-free nitrogen atmosphere andthe mixture is maintained under gentle reflux. 1,2-Dibromoethane (0.22ml, 2.60 mmol) is added dropwise over a period of 1/2 hour. The mixtureis heated gently until the magnesium is nearly consumed (ca. 1/2 hour).2-(p-Chlorobenzyl)-6-methoxy-3-keto-(2H)benzothiophene (792 mg, 2.60mmol) in THF (50 ml) is added dropwise to the mixture at 0° C. Thereaction mixture is allowed to warm to room temperature and refluxed for18 hours. The cooled mixture is quenched with saturated ammoniumchloride (30 ml), extracted with dichloromethane (3×40 ml), washed withwater, dried (anhydrous sodium sulphate) and the solvent evaporated off.The remaining material is dehydrated with concentrated hydrochloric acid(2 ml) in ethanol (25 ml). The mixture is refluxed for 2 hours,whereupon ethanol is removed. It is then basified with 10% sodiumhydroxide (20 ml) and extracted with dichloromethane (3×40 ml). Thedichloromethane extract is washed with water (3×20 ml), dried (anhydroussodium sulphate) and the solvent is evaporated. Elution with(dichloromethane:hexane:triethylamine=1:2:0.3), yields a crude titledproduct which is recrystallized from methanol to give titled product.Dry hydrogen chloride gas is bubbled through an ethereal solution of thefree base for 10 minutes and cooled to 0° C. The hydrochloride saltwhich precipitated out is filtered off at the pump, rinsed with ether,and recrystallized from isopropanol to yield the hydrochloride salt.

EXAMPLE 32-(p-Chlorobenzyl)-3-[p-(2-(1-piperidino)ethoxy)phenyl]-6-methoxybenzo[b]thiophene

n-Butyl lithium (5.27 ml, 7.80 mM) in THF is added dropwise to asolution of 4-[2-(1-piperidino)ethoxy]phenyl bromide (1.45 g, 7.80 mmol)in THF (5 ml) at -78° C. After stirring for 30 minutes at -78° C.,2-(p-chlorobenzyl)-6-methoxy-3-keto-(2H)benzothiophene (1.58 g, 5.20mmol) in THF is added dropwise to the mixture at -78° C. The reactionmixture is stirred at -78° C. for 1 hour. It is warmed to roomtemperature. After stirring for 18 hours, it is quenched with saturatedammonium chloride and THF is evaporated off. The residue is dissolved inethyl acetate (30 ml), washed with sodium chloride (2×20 ml), dried(anhydrous magnesium sulphate) and the solvent evaporated off.Thereafter, the material is purified by flash chromatography (eluent,dichloromethane:hexane:triethylamine=1:1:0.2), and a solution ofmethanol (0.40 ml) saturated with dry hydrogen chloride gas is added tothe material dissolved in sodium-dried ether (40 ml). The reactionmixture is stirred at room temperature for 1/2 hour. A precipitate isformed which is filtered and recrystallized from ethanol to give thehydrochloride salt of the title compound.

EXAMPLE 42-(p-Chlorobenzyl)-3-[p-(2-(1-morpholino)ethoxy)phenyl-6-methoxybenzo[b]thiophene

(a) n-Butyl lithium (1.41 ml of a 1.56M solution, 2.2 mmol) is addeddropwise to a solution of 4-[2-(1-morpholino)ethoxy]phenyl bromide (630mg, 2.2 mmol) in THF (5 ml) at -78° C. After stirring for 30 minutes at-78° C., 2-(p-chlorobenzyl)-6-methoxy-3-keto-(2H)benzothiophene (609 mg,2.0 mmol) in THF (2 ml) is added dropwise to the mixture at -78° C. Thereaction mixture is allowed to warm to room temperature. After stirringfor 18 hours, it is quenched with saturated ammonium chloride, THF isevaporated off to yield an alcohol which is dehydrated with concentratedhydrochloric acid (2 ml) in ethanol (10 ml). The mixture is refluxed for2 hours, whereupon ethanol is removed. It is then basified with 10%sodium hydroxide (20 ml) and extracted with dichloromethane (3×40 ml).

The dichloromethane extract is washed with water (3×20 ml), dried(anhydrous sodium sulphate) and the solvent is evaporated off. Thematerial is then eluted withdichromethane:hexane:triethylamine=1:2:0.3), and cooled to 0° C. Dryhydrogen chloride gas is bubbled for 10 minutes. The hydrochloride saltof the title compound is obtained after recrystallized from isopropanol.

(b) Magnesium bromide etherate (0.194 g, 0.75 mmol) is dissolved in 20ml ether in a two-necked flask under oxygen-free nitrogen atmosphere atroom temperature. After stirring for 1 hour,2-(p-chlorophenylmethylene)-3-[p-(2-(1-morpholino)ethoxy)phenyl]-6-methoxy-2,3-dihydrobenzo[b]thiophen-3-ol(0.255 g, 0.5 mmol) in ether (10 ml) is added dropwise to the mixture.After stirring for 4 hours, LiAlH₄ (0.076 g, 2.0 mmol) is added to theabove reaction mixture. The reaction mixture is left overnight at roomtemperature. After stirring overnight, ethyl acetate is added dropwisetill hydrogen evolution ceased. The above mixture is washed with waterfollowed by brine 2×20 ml), dried (anhydrous magnesium sulfate) and thesolvent is evaporated off, followed by chromatography on silica. Elutionwith (chloroform:hexane:triethylamine=1:1:0.2) yields titled product.

EXAMPLE 52-(p-Chorophenylmethylene)-3-[p-(2-(1-morpholino)ethoxy)phenyl]-6-methoxy-2,3-dihydrobenzo[b]thiophen-3-ol

n-Butyl lithium (2.87 mi of a 1.28M solution, 3.68 mmol) is addeddropwise to solution of 4-[2-(1-morpholino)ethoxy]phenyl bromide (0.80ml, 3.67 mmol) in THF (10 ml) at -78° C. under nitrogen atmosphere.After stirring for 1 hour at -78° C.,2-(p-chlorobenzylidene)-6-methoxy-3-keto-benzothiophene (1.06 g, 3.50mmol) in THF is added dropwise to the mixture at -78° C. The reactionmixture is left overnight at room temperature. It is quenched withsaturated ammonium chloride, THF is evaporated off, followed byextracting with ethyl acetate (2×20 ml). The ethyl acetate extract iswashed with water followed by brine (2×20 ml), dried (anhydrousmagnesium sulfate) and the solvent is evaporated, followed bychromatography on silica gel. Elution with(chloroform:hexane:triethylamine=1:1:0.2) yields titled product.

EXAMPLE 62-(p-Fluorobenzyl)-3-[p-(2-diethylaminoethoxy)phenyl]-6-methoxybenzo[b]thiophene

The methods of Example 5 followed by Example 4b are used for thepreparation of2-(p-fluorobenzyl)-3-[p-(2-diethylaminoethoxy)phenyl]-6-methoxybenzo[b]thiophene.The specific gram proportions of reactants used are similar and are notmeant to be limiting and can be varied around idealized stoichiometricproportion in the usual way known to those skilled in the art.

EXAMPLE 72-(p-Fluorobenzyl)-3-[p-(2-(1-pyrollidino)ethoxy)phenyl]-6-methoxybenzo[b]thiophene

The methods of Example 5 followed by Example 4b are used for thepreparation of2-(p-fluorobenzyl)-3-[p-(2-(1-pyrollidino)ethoxy)phenyl]-6-methoxybenzo[b]thiophene.

EXAMPLE 82-(p-Fluorobenzyl)-3-[p-(2-(1-piperidino)ethoxy)phenyl]-6-methoxybenzo[b]thiophene

The methods of Example 5 followed by Example 4b are used for thepreparation of2-(p-fluorobenzyl)-3-[p-(2-(1-piperidino)ethoxy)phenyl]-6-methoxybenzo[b]thiophene.

EXAMPLE 92-(p-Fluorobenzyl)-3-[p-(2-(1-morpholino)ethoxy)phenyl]-6-methoxybenzo[b]thiophene

The methods of Example 5 followed by Example 4b are used for thepreparation of2-(p-fluorobenzyl)-3-[p-(2-(1-morpholino)ethoxy)phenyl]-6-methoxybenzo[b]thiophene.

EXAMPLE 10 2-(p-Methoxyphenyl)-3-[p-(2-(1-piperidenyl)ethoxy)phenyl]-6-methoxy benz[b]thiophene

The methods of Example 5 followed by Example 4b are used for thepreparation of 2-(p-Methoxyphenyl)-3-[p-(2-(1-piperidenyl)ethoxy)phenyl]-6-methoxy benz[b]thiophene.

Although the free-base form of formula I compounds can be used in themethods of the present invention, it may be preferred to prepare and usea pharmaceutically acceptable salt form. Thus, the compounds used in themethods of this invention primarily form pharmaceutically acceptableacid addition salts with a wide variety of organic and inorganic acids,and include the physiologically acceptable salts which are often used inpharmaceutical chemistry. Such salts are also part of this invention.Typical inorganic acids used to form such salts include hydrochloric,hydrobromic, hydroiodic, nitric, sulfuric, phosphoric, hypophosphoric,and the like. Salts derived from organic acids, such as aliphatic monoand dicarboxylic acids, phenyl substituted alkanoic acids,hydroxyalkanoic and hydroxyalkandioic acids, aromatic acids, aliphaticand aromatic sulfonic acids, may also be used. Such pharmaceuticallyacceptable salts thus include acetate, phenylacetate, trifluoroacetate,acrylate, ascorbate, benzoate, chlorobenzoate, dinitrobenzoate,hydroxybenzoate, methoxybenzoate, methylbenzoate, o-acetoxybenzoate,naphthalene-2-benzoate, bromide, isobutyrate, phenylbutyrate,β-hydroxybutyrate, butyne-1,4-dioate, hexyne-1,4-dioate, caprate,caprylate, chloride, cinnamate, citrate, formate, fumarate, glycollate,heptanoate, hippurate, lactate, malate, maleate, hydroxymaleate,malonate, mandelate, mesylate, nicotinate, isonicotinate, nitrate,oxalate, phthalate, terephthalate, phosphate, monohydrogenphosphate,dihydrogenphosphate, metaphosphate, pyrophosphate, propiolate,propionate, phenylpropionate, salicylate, sebacate, succinate, suberate,sulfate, bisulfate, pyrosulfate, sulfite, bisulfite, sulfonate,benzenesulfonate, p-bromophenylsulfonate, chlorobenzenesulfonate,ethanesulfonate, 2-hydroxyethanesulfonate, methanesulfonate,naphthalene-1-sulfonate, naphthalene-2-sulfonate, p-toluenesulfonate,xylenesulfonate, tartarate, and the like. A preferred salt is thehydrochloride salt.

The pharmaceutically acceptable acid addition salts are typically formedby reacting a compound of formula I with an equimolar or excess amountof acid. The reactants are generally combined in a mutual solvent suchas diethyl ether or ethyl acetate. The salt normally precipitates out ofsolution within about one hour to 10 days and can be isolated byfiltration or the solvent can be stripped off by conventional means.

The pharmaceutically acceptable salts generally have enhanced solubilitycharacteristics compared to the compound from which they are derived,and thus are often more amenable to formulation as liquids or emulsions.

The following examples are presented to further illustrate thepreparation of compounds of the present invention. It is not intendedthat the invention be limited in scope by reason of any of the followingexamples.

TEST PROCEDURE General Preparation Procedure

In the examples illustrating the methods, a post-menopausal model isused in which effects of different treatments upon circulating lipidswere determined.

Seventy-five day old female Sprague Dawley rats (weight range of 200 to225 g) are obtained from Charles River Laboratories (Portage, Mich.).The animals are either bilaterally ovariectomized (OVX) or exposed to aSham surgical procedure at Charles River Laboratories, and then shippedafter one week. Upon arrival, they are housed in metal hanging cages ingroups of 3 or 4 per cage and had ad libitum access to food (calciumcontent approximately 0.5%) and water for one week. Room temperature ismaintained at 22.2°±1.7° C. with a minimum relative humidity of 40%. Thephotoperiod in the room is 12 hours light and 12 hours dark.

Dosing Regimen Tissue Collection

After a one week acclimation period (therefore, two weeks post-OVX)daily dosing with test compound is initiated. 17°-ethynyl estradiol orthe test compound are given orally, unless otherwise stated, as asuspension in 1% carboxymethylcellulose or dissolved in 20%cyclodextrin. Animals are dosed daily for 4 days. Following the dosingregimen, animals are weighed and anesthetized with a ketamine: Xylazine(2:1, V:V) mixture and a blood sample is collected by cardiac puncture.The animals are then sacrificed by asphyxiation with CO₂, the uterus isremoved through a midline incision, and a wet uterine weight isdetermined.

Cholesterol Analysis

Blood samples are allowed to clot at room temperature for 2 hours, andserum is obtained following centrifugation for 10 minutes at 3000 rpm.Serum cholesterol is determined using a Boehringer Mannheim Diagnosticshigh performance cholesterol assay. Briefly the cholesterol is oxidizedto cholest-4-en-3-one and hydrogen peroxide. The hydrogen peroxide isthen reacted with phenol and 4-aminophenazone in the presence ofperoxidase to produce a p-quinone imine dye, which is readspectrophotemetrically at 500 nm. Cholesterol concentration is thencalculated against a standard curve. The entire assay is automated usinga Biomek Automated Workstation.

Uterine Eosinophil Peroxidase (EPO) Assay

Uteri are kept at 4° C. until time of enzymatic analysis. The uteri arethen homogenized in 50 volumes of 50 mM Tris buffer (pH--8.0 containing0.005% Triton X-100. Upon addition of 0.01% hydrogen peroxide and 10 mMO-phenylenediamine (final concentrations) in Tris buffer, increase inabsorbance is monitored for one minute at 450 nm. The presence ofeosonophils in the uterus is an indication of estrogenic activity of acompound. The maximal velocity of a 15 second interval is determinedover the initial, linear portion of the reaction curve.

Source of Compound

17α-ethynyl estradiol is obtained from Sigma Chemical Co., St. Louis,Mo.

Osteoporosis Test Procedure

Following the General Preparation Procedure, infra, the rats are treateddaily for 35 days (6 rats per treatment group) and sacrificed by carbondioxide asphyxiation on the 36th day. The 35 day time period issufficient to allow maximal reduction in bone density, measured asdescribed herein. At the time of sacrifice, the uteri are removed,dissected free of extraneous tissue, and the fluid contents are expelledbefore determination of wet weight in order to confirm estrogendeficiency associated with complete ovariectomy. Uterine weight isroutinely reduced about 75% in response to ovariectomy. The uteri arethen placed in 10% neutral buffered formalin to allow for subsequenthistological analysis.

The right femurs are excised and digitilized x-rays generated andanalyzed by an image analysis program (NIH image) at the distalmetaphysis. The proximal aspect of the tibiae from these animals arealso scanned by quantitative computed tomography.

In accordance with the above procedures, compounds of the presentinvention and ethynyl estradiol (EE₂) in 20% hydroxypropylβ-cyclodextrin are orally administered to test animals.

MCF-7 Proliferation Assay

MCF-7 breast adenocarcinoma cells (ATCC HTB 22) are maintained in MEM(minimal essential medium, phenol red-free, Sigma, St. Louis, Mo.)supplemented with 10% fetal bovine serum (FBS) (V/V), L-glutamine (2mM), sodium pyruvate (1 mM), HEPES{(N-[2-hydroxyethyl]piperazine-N'-[2-ethanesulfonic acid]10 mM},non-essential amino acids and bovine insulin (1 ug/mL) (maintenancemedium). Ten days prior to assay, MCF-7 cells are switched tomaintenance medium supplemented with 10% dextran coated charcoalstripped fetal bovine serum (DCC-FBS) assay medium) in place of 10% FBSto deplete internal stores of steroids. MCF-7 cells are removed frommaintenance flasks using cell dissociation medium (Ca++/Mg++ free HBSS(phenol red-free) supplemented with 10 mM HEPES and 2 mM EDTA). Cellsare washed twice with assay medium and adjusted to 80,000 cells/mL.Approximately 100 μL (8,000 cells) are added to flat-bottom microculturewells (Costar 3596) and incubated at 37° C. in a 5% CO₂ humidifiedincubator for 48 hours to allow for cell adherence and equilibrationafter transfer. Serial dilutions of drugs or DMSO as a diluent controlare prepared in assay medium and 50 μL transferred to triplicatemicrocultures followed by 50 μL assay medium for a final volume of 200μL. After an additional 48 hours at 37° C. in a 5% CO₂ humidifiedincubator, microcultures are pulsed with tritiated thymidine (1uCi/well) for 4 hours. Cultures are terminated by freezing at -70° C.for 24 hours followed by thawing and harvesting of microcultures using aSkatron Semiautomatic Cell Harvester. Samples are counted by liquidscintillation using a Wallac BetaPlace β counter.

DMBA-Induced Mammary Tumor Inhibition

Estrogen-dependent mammary tumors are produced in female Sprague-Dawleyrats which are purchased from Harlan Industries, Indianapolis, Ind. Atabout 55 days of age, the rats receive a single oral feeding of 20 mg of7,12-dimethylbenz[a]anthracene (DMBA). About 6 weeks after DMBAadministration, the mammary glands are palpated at weekly intervals forthe appearance of tumors. Whenever one or more tumors appear, thelongest and shortest diameters of each tumor are measured with a metriccaliper, the measurements are recorded, and that animal is selected forexperimentation. An attempt is made to uniformly distribute the varioussizes of tumors in the treated and control groups such thataverage-sized tumors are equivalently distributed between test groups.Control groups and test groups for each experiment contain 5 to 9animals.

Compounds of Formula I are administered either through intraperitonealinjections in 2% acacia, or orally. Orally administered compounds areeither dissolved or suspended in 0.2 mL corn oil. Each treatment,including acacia and corn oil control treatments, is administered oncedaily to each test animal. Following the initial tumor measurement andselection of test animals, tumors are measured each week by theabove-mentioned method. The treatment and measurements of animalscontinue for 3 to 5 weeks at which time the final areas of the tumorsare determined. For each compound and control treatment, the change inthe mean tumor area is determined.

Uterine Fibrosis Test Procedures

Assay 1

Between 3 and 20 women having uterine fibrosis are administered acompound of the present invention. The amount of compound administeredis from 0.1 to 1000 mg/day, and the period of administration is 3months.

The women are observed during the period of administration, and up to 3months after discontinuance of administration, for effects on uterinefibrosis.

Assay 2

The same procedure is used as in Test 1, except the period ofadministration is 6 months.

Assay 3

The same procedure is used as in Test 1, except the period ofadministration is 1 year.

Assay 4

A. Induction of Fibroid Tumors in Guinea Pig

Prolonged estrogen stimulation is used to induce leiomyomata in sexuallymature female guinea pigs. Animals are dosed with estradiol 3-5 timesper week by injection for 2-4 months or until tumors arise. Treatmentsconsisting of a compound of the invention or vehicle is administereddaily for 3-16 weeks and then animals are sacrificed and the uteriharvested and analyzed for tumor regression.

B. Implantation of Human Uterine Fibroid Tissue in Nude Mice

Tissue from human leiomyomas are implanted into the peritoneal cavityand or uterine myometrium of sexually mature, castrated, female, nudemice. Exogenous estrogen are supplied to induce growth of the explantedtissue. In some cases, the harvested tumor cells are cultured in vitroprior to implantation. Treatment consisting of a compound of the presentinvention or vehicle is supplied by gastric lavage on a daily basis for3-16 weeks and implants are removed and measured for growth orregression. At the time of sacrifice, the uteri is harvested to assessthe status of the organ.

Assay 5

A. Tissue from human uterine fibroid tumors is harvested and maintained,in vitro, as primary nontransformed cultures. Surgical specimens arepushed through a sterile mesh or sieve, or alternately teased apart fromsurrounding tissue to produce a single cell suspension. Cells aremaintained in media containing 10% serum and antibiotic. Rates of growthin the presence and absence of estrogen are determined. Cells areassayed for their ability to produce complement component C3 and theirresponse to growth factors and growth hormone. In vitro cultures areassessed for their proliferative response following treatment withprogestins, GnRH, a compound of the present invention and vehicle.Levels of steroid hormone receptors are assessed weekly to determinewhether important cell characteristics are maintained in vitro. Tissuefrom 5-25 patients are utilized.

Activity in at least one of the above tests indicates the compounds ofthe present invention are of potential in the treatment of uterinefibrosis.

Endometriosis Test Procedure

In Tests 1 and 2, effects of 14-day and 21-day administration ofcompounds of the present invention on the growth of explantedendometrial tissue can be examined.

Assay 1

Twelve to thirty adult CD strain female rats are used as test animals.They are divided into three groups of equal numbers. The estrous cycleof all animals is monitored. On the day of proestrus, surgery isperformed on each female. Females in each group have the left uterinehorn removed, sectioned into small squares, and the squares are looselysutured at various sites adjacent to the mesenteric blood flow. Inaddition, females in Group 2 have the ovaries removed.

On the day following surgery, animals in Groups 1 and 2 receiveintraperitoneal injections of water for 14 days whereas animals in Group3 receive intraperitoneal injections of 1.0 mg of a compound of thepresent invention per kilogram of body weight for the same duration.Following 14 days of treatment, each female is sacrificed and theendometrial explants, adrenals, remaining uterus, and ovaries, whereapplicable, are removed and prepared for histological examination. Theovaries and adrenals are weighed.

Assay 2

Twelve to thirty adult CD strain female rats are used as test animals.They are divided into two equal groups. The estrous cycle of all animalsis monitored. On the day of proestrus, surgery is performed on eachfemale. Females in each group have the left uterine horn removed,sectioned into small squares, and the squares are loosely sutured atvarious sites adjacent to the mesenteric blood flow.

Approximately 50 days following surgery, animals assigned to Group 1receive intraperitoneal injections of water for 21 days whereas animalsin Group 2 receive intraperitoneal injections of 1.0 mg of a compound ofthe present invention per kilogram of body weight for the same duration.Following 21 days of treatment, each female is sacrificed and theendometrial explants and adrenals are removed and weighed. The explantsare measured as an indication of growth. Estrous cycles are monitored.

Assay 3

A. Surgical Induction of Endometriosis

Autographs of endometrial tissue are used to induce endometriosis inrats and/or rabbits. Female animals at reproductive maturity undergobilateral oophorectomy, and estrogen is supplied exogenously thusproviding a specific and constant level of hormone. Autologousendometrial tissue is implanted in the peritoneum of 5-150 animals andestrogen supplied to induce growth of the explanted tissue. Treatmentconsisting of a compound of the present invention is supplied by gastriclavage on a daily basis for 3-16 weeks, and implants are removed andmeasured for growth or regression. At the time of sacrifice, the intacthorn of the uterus is harvested to assess status of endometrium.

B. Implantation of Human Endometrial Tissue in Nude Mice

Tissue from human endometrial lesions is implanted into the peritoneumof sexually mature, castrated, female, nude mice. Exogenous estrogen issupplied to induce growth of the explanted tissue. In some cases, theharvested endometrial cells are cultured in vitro prior to implantation.Treatment consisting of a compound of the present invention supplied bygastric lavage on a daily basis for 3-16 weeks, and implants are removedand measured for growth or regression. At the time of sacrifice, theuteri is harvested to assess the status of the intact endometrium.

Assay 4

A. Tissue from human endometrial lesions is harvested and maintained invitro as primary nontransformed cultures. Surgical specimens are pushedthrough a sterile mesh or sieve, or alternately teased apart fromsurrounding tissue to produce a single cell suspension. Cells aremaintained in media containing 10% serum and antibiotic. Rates of growthin the presence and absence of estrogen are determined. Cells areassayed for their ability to produce complement component C3 and theirresponse to growth factors and growth hormone. In vitro cultures areassessed for their proliferative response following treatment withprogestins, GnRH, a compound of the invention, and vehicle. Levels ofsteroid hormone receptors are assessed weekly to determine whetherimportant cell characteristics are maintained in vitro. Tissue from 5-25patients is utilized.

Activity in any of the above assays indicates that the compounds of thepresent invention are useful in the treatment of endometriosis.

Inhibition of Aortal Smooth Cell Proliferation/Restenosis Test Procedure

Compounds of the present invention have capacity to inhibit aortalsmooth cell proliferation. This can be demonstrated by using culturedsmooth cells derived from rabbit aorta, proliferation being determinedby the measurement of DNA synthesis. Cells are obtained by explantmethod as described in Ross, J. of Cell Bio. 50: 172 (1971). Cells areplated in 96 well microtiter plates for five days. The cultures becomeconfluent and growth arrested. The cells are then transferred toDulbecco's Modified Eagle's Medium (DMEM) containing 0.5-2% plateletpoor plasma, 2 mM L-glutamine, 100 U/ml penicillin, 100 mg mlstreptomycin, 1 mC/ml ³ H-thymidine, 20 ng/ml platelet-derived growthfactor, and varying concentrations of the present compounds. Stocksolution of the compounds is prepared in dimethyl sulphoxide and thendiluted to appropriate concentration (0.01-30 mM) in the above assaymedium. Cells are then incubated at 37° C. for 24 hours under 5% CO₂/95% air. At the end of 24 hours, the cells are fixed in methanol. ³ Hthymidine incorporation in DNA is then determined by scintillationcounting as described in Bonin, et al., Exp. Cell Res, 181: 475-482(1989).

Inhibition of aortal smooth muscle cell proliferation by the compoundsof the present invention are further demonstrated by determining theireffects on exponentially growing cells. Smooth muscle cells from rabbitaortae are seeded in 12 well tissue culture plates in DMEM containing10% fetal bovine serum, 2 mM L-glutamine, 100 U/ml penicillin, and 100mg/ml streptomycin. After 24 hours, the cells are attached and themedium is replaced with DMEM containing 10% serum, 2 mM L-glutamine, 100U/ml penicillin, 100 mg/ml streptomycin, and desired concentrations ofthe compounds. Cells are allowed to grow for four days. Cells aretreated with trypsin and the number of cells in each culture isdetermined by counting using a ZM-Coulter counter.

Activity in the above assays indicates that the compounds of the presentinvention are of potential in the treatment of restenosis.

The present invention also provides a method of alleviatingpost-menopausal syndrome in women which comprises the aforementionedmethod using compounds of Formula I and further comprises administeringto a woman an effective amount of estrogen or progestin. Thesetreatments are particularly useful for treating osteoporosis andlowering serum cholesterol because the patient will receive the benefitsof each pharmaceutical agent while the compounds of the presentinvention would inhibit undesirable side-effects of estrogen andprogestin. Activity of these combination treatments in any of thepost-menopausal tests, infra, indicates that the combination treatmentsare useful for alleviating the symptoms of post-menopausal symptoms inwomen.

Various forms of estrogen and progestin are commercially available.Estrogen-based agents include, for example, ethynyl estrogen (0.01-0.03mg/day), mestranol (0.05-0.15 mg/day), and conjugated estrogenichormones such as Premarin® (Wyeth-Ayerst; 0.3-2.5 mg/day).Progestin-based agents include, for example, medroxyprogesterone such asProvera® (Upjohn; 2.5-10 mg/day), norethylnodrel (1.0-10.0 mg/day), andnonethindrone (0.5-2.0 mg/day). A preferred estrogen-based compound isPremarin, and norethylnodrel and norethindrone are preferredprogestin-based agents.

The method of administration of each estrogen- and progestin-based agentis consistent with that which is known in the art. For the majority ofthe methods of the present invention, compounds of Formula I areadministered continuously, from 1 to 3 times daily. However, cyclicaltherapy may especially be useful in the treatment of endometriosis ormay be used acutely during painful attacks of the disease. In the caseof restenosis, therapy may be limited to short (1-6 months) intervalsfollowing medical procedures such as angioplasty.

As used herein, the term "effective amount" means an amount of compoundof the present invention which is capable of alleviating the symptoms ofthe various pathological conditions herein described. The specific doseof a compound administered according to this invention will, of course,be determined by the particular circumstances surrounding the caseincluding, for example, the compound administered, the route ofadministration, the state of being of the patient, and the pathologicalcondition being treated. A typical daily dose will contain a nontoxicdosage level of from about 5 mg to about 600 mg/day of a compound of thepresent invention. Preferred daily doses generally will be from about 15mg to about 80 mg/day.

The compounds of this invention can be administered by a variety ofroutes including oral, rectal, transdermal, subcutaneous, intravenous,intramuscular, and intranasal. These compounds preferably are formulatedprior to administration, the selection of which will be decided by theattending physician. Thus, another aspect of the present invention is apharmaceutical composition comprising an effective amount of a compoundof Formula I, or a pharmaceutically acceptable salt thereof, optionallycontaining an effective amount of estrogen or progestin, and apharmaceutically acceptable carrier, diluent, or excipient.

The total active ingredients in such formulations comprises from 0.1% to99.9% by weight of the formulation. By "pharmaceutically acceptable" itis meant the carrier, diluent, excipients and salt must be compatiblewith the other ingredients of the formulation, and not deleterious tothe recipient thereof.

Pharmaceutical formulations of the present invention can be prepared byprocedures known in the art using well known and readily availableingredients. For example, the compounds of formula I, with or without anestrogen or progestin compound, can be formulated with commonexcipients, diluents, or carriers, and formed into tablets, capsules,suspensions, powders, and the like. Examples of excipients, diluents,and carriers that are suitable for such formulations include thefollowing: fillers and extenders such as starch, sugars, mannitol, andsilicic derivatives; binding agents such as carboxymethyl cellulose andother cellulose derivatives, alginates, gelatin, andpolyvinyl-pyrrolidone; moisturizing agents such as glycerol;disintegrating agents such as calcium carbonate and sodium bicarbonate;agents for retarding dissolution such as paraffin; resorptionaccelerators such as quaternary ammonium compounds; surface activeagents such as cetyl alcohol, glycerol monostearate; adsorptive carrierssuch as kaolin and bentonite; and lubricants such as talc, calcium andmagnesium stearate, and solid polyethyl glycols.

The compounds also can be formulated as elixirs or solutions forconvenient oral administration or as solutions appropriate forparenteral administration, for example, by intramuscular, subcutaneousor intravenous routes. Additionally, the compounds are well suited toformulation as sustained release dosage forms and the like. Theformulations can be so constituted that they release the activeingredient only or preferably in a particular physiological location,possibly over a period of time. The coatings, envelopes, and protectivematrices may be made, for example, from polymeric substances or waxes.

Compounds of formula I, alone or in combination with a pharmaceuticalagent of the present invention, generally will be administered in aconvenient formulation. The following formulation examples only areillustrative and are not intended to limit the scope of the presentinvention.

Formulations

In the formulations which follow, "active ingredient" means a compoundof formula I, or a salt or solvate thereof.

    ______________________________________                                        Formulation 1: Gelatin Capsules                                               Hard gelatin capsules are prepared using the following:                       Ingredient        Quantity (mg/capsule)                                       ______________________________________                                        Active ingredient   0.1-1000                                                  Starch, NF         0-650                                                      Starch flowable powder                                                                           0-650                                                      Silicone fluid 350 centistokes                                                                   0-15                                                       ______________________________________                                    

The formulation above may be changed in compliance with the reasonablevariations provided.

A tablet formulation is prepared using the ingredients below:

    ______________________________________                                        Formulation 2: Tablets                                                        Ingredient        Quantity (mg/tablet)                                        ______________________________________                                        Active ingredient   2.5-1000                                                  Cellulose, microcrystalline                                                                     200-650                                                     Silicon dioxide, fumed                                                                           10-650                                                     Stearate acid      5-15                                                       ______________________________________                                    

The components are blended and compressed to form tablets.

Alternatively, tablets each containing 2.5-1000 mg of active ingredientare made up as follows:

    ______________________________________                                        Formulation 3-: Tablets                                                       Ingredient          Quantity (mg/tablet)                                      ______________________________________                                        Active ingredient   25-1000                                                   Starch              45                                                        Cellulose, microcrystalline                                                                       35                                                        Polyvinylpyrrolidone                                                                              4                                                         (as 10% solution in water)                                                    Sodium carboxymethyl cellulose                                                                    4.5                                                       Magnesium stearate  0.5                                                       Talc                1                                                         ______________________________________                                    

The active ingredient, starch, and cellulose are passed through a No. 45mesh U.S. sieve and mixed thoroughly. The solution ofpolyvinylpyrrolidone is mixed with the resultant powders which are thenpassed through a No. 14 mesh U.S. sieve. The granules so produced aredried at 50°-60° C. and passed through a No. 18 mesh U.S. sieve. Thesodium carboxymethyl starch, magnesium stearate, and talc, previouslypassed through a No. 60 U.S. sieve, are then added to the granuleswhich, after mixing, are compressed on a tablet machine to yieldtablets.

Suspensions each containing 0.1-1000 mg of medicament per 5 ml dose aremade as follows:

    ______________________________________                                        Formulation 4: Suspensions                                                    Ingredient             Quantity (mg/5 ml)                                     ______________________________________                                        Active ingredient      0.1-1000  mg                                           Sodium carboxymethyl cellulose                                                                       50        mg                                           Syrup                  1.25      mg                                           Benzoic acid solution  0.10      mL                                           Flavor                 q.v.                                                   Color                  q.v.                                                   Purified water to      5         mL                                           ______________________________________                                    

The medicament is passed through a No. 45 mesh U.S. sieve and mixed withthe sodium carboxymethyl cellulose and syrup to form a smooth paste. Thebenzoic acid solution, flavor, and color are diluted with some of thewater and added, with stirring. Sufficient water is then added toproduce the required volume.

An aerosol solution is prepared containing the following ingredients:

    ______________________________________                                        Formulation 5: Aerosol                                                        Ingredient           Quantity (% by weight)                                   ______________________________________                                        Active ingredient    0.25                                                     Ethanol              25.75                                                    Propellant 22 (Chlorodifluoromethane)                                                              70.00                                                    ______________________________________                                    

The active ingredient is mixed with ethanol and the mixture added to aportion of the propellant 22, cooled to 30° C., and transferred to afilling device. The required amount is then fed to a stainless steelcontainer and diluted with the remaining propellant. The valve units arethen fitted to the container.

Suppositories are prepared as follows:

    ______________________________________                                        Formulation 6: Suppositories                                                  Ingredient       Quantity (mg/suppository)                                    ______________________________________                                        Active ingredient                                                                              250                                                          Saturated fatty acid glycerides                                                                2,000                                                        ______________________________________                                    

The active ingredient is passed through a No. 60 mesh U.S. sieve andsuspended in the saturated fatty acid glycerides previously melted usingthe minimal necessary heat. The mixture is then poured into asuppository mold of nominal 2 g capacity and allowed to cool.

An intravenous formulation is prepared as follows:

    ______________________________________                                        Formulation 7: Intravenous Solution                                           Ingredient            Quantity                                                ______________________________________                                        Active ingredient     50     mg                                               Isotonic saline       1,000  mL                                               ______________________________________                                    

The solution of the above ingredients is intravenously administered to apatient at a rate of about 1 mL per minute.

    ______________________________________                                        Formulation 8: Combination Capsule I                                          Ingredient     Quantity (mg/capsule)                                          ______________________________________                                        Active ingredient                                                                            50                                                             Premarin       1                                                              Avicel pH 101  50                                                             Starch 1500    117.50                                                         Silicon Oil    2                                                              Tween 80       0.50                                                           Cab-O-Sil      0.25                                                           ______________________________________                                    

    ______________________________________                                        Formulation 9: Combination Capsule II                                         Ingredient     Quantity (mg/capsule)                                          ______________________________________                                        Active ingredient                                                                            50                                                             Norethylnodrel 5                                                              Avicel pH 101  82.50                                                          Starch 1500    90                                                             Silicon Oil    2                                                              Tween 80       0.50                                                           ______________________________________                                    

    ______________________________________                                        Formulation 10: Combination Tablet                                            Ingredient      Quantity (mg/capsule)                                         ______________________________________                                        Active ingredient                                                                             50                                                            Premarin        1                                                             Corn Starch NF  50                                                            Povidone, K29-32                                                                              6                                                             Avicel pH 101   41.50                                                         Avicel pH 102   136.50                                                        Crospovidone XL10                                                                             2.50                                                          Magnesium Stearate                                                                            0.50                                                          Cab-O-Sil       0.50                                                          ______________________________________                                    

We claim:
 1. A method for alleviating the symptoms of post-menopausalsyndrome comprising administering to a woman in need of such treatmentan effective amount of a compound of formula I ##STR21## wherein R₁ ishydrogen, a hydroxy protecting group, or methyl;R₂ and R₃ areindependently C₁ -C₆ alkyl, or R₂ and R₃, together with the nitrogen towhich they are attached, form a heterocycle selected from the groupconsisting of pyrrolidino, hexamethyleneimino, morpholino, andpiperidino; Z is bromo, chloro, fluoro, hydroxy, protected hydroxy, orhydrogen; and pharmaceutically acceptable salts and solvates thereof. 2.A method according to claim 1 wherein the post-menopausal syndromepathological condition is osteoporosis.
 3. A method according to claim 1wherein the post-menopausal syndrome pathological condition is relatedto a cardiovascular disease.
 4. A method according to claim 3 whereinthe cardiovascular disease is hyperlipidemia.
 5. A method for inhibitinguterine fibroid disease comprising administering to a woman in need ofsuch treatment an effective amount of a compound of formula I ##STR22##wherein R₁ is hydrogen, a hydroxy protecting group, or methyl;R₂ and R₃are independently C₁ -C₆ alkyl, or R₂ and R₃, together with the nitrogento which they are attached, form a heterocycle selected from the groupconsisting of pyrrolidino, hexamethyleneimino, morpholino, andpiperidino; Z is bromo, chloro, fluoro, hydroxy, protected hydroxy, orhydrogen; and pharmaceutically acceptable salts and solvates thereof. 6.A method for inhibiting endometriosis comprising administering to awoman in need of such treatment an effective amount of a compound offormula I ##STR23## wherein R₁ is hydrogen, a hydroxy protecting group,or methyl;R₂ and R₃ are independently C₁ -C₆ alkyl, or R₂ and R₃,together with the nitrogen to which they are attached, form aheterocycle selected from the group consisting of pyrrolidino,hexamethyleneimino, morpholino, and piperidino; Z is bromo, chloro,fluoro, hydroxy, protected hydroxy, or hydrogen; and pharmaceuticallyacceptable salts and solvates thereof.
 7. A method for inhibiting aortalsmooth muscle cell proliferation comprising administering to a human inneed of such treatment an effective amount of a compound of formula I##STR24## wherein R₁ is hydrogen, a hydroxy protecting group, ormethyl;R₂ and R₃ are independently C₁ -C₆ alkyl, or R₂ and R₃, togetherwith the nitrogen to which they are attached, form a heterocycleselected from the group consisting of pyrrolidino, hexamethyleneimino,morpholino, and piperidino; Z is bromo, chloro, fluoro, hydroxy,protected hydroxy, or hydrogen; and pharmaceutically acceptable saltsand solvates thereof.
 8. A method for inhibiting restenosis comprisingadministering to a human in need of such treatment an effective amountof a compound of formula I ##STR25## wherein R₁ is hydrogen, a hydroxyprotecting group, or methyl;R₂ and R₃ are independently C₁ -C₆ alkyl, orR₂ and R₃, together with the nitrogen to which they are attached, form aheterocycle selected from the group consisting of pyrrolidino,hexamethyleneimino, morpholino, and piperidino; Z is bromo, chloro,fluoro, hydroxy, protected hydroxy, or hydrogen; and pharmaceuticallyacceptable salts and solvates thereof.